Analysis of mRNA is very important to understand various biological phenomena. Based on the founding of RNA-dependent DNA polymerase, i.e., reverse transcriptase, in retroviruses, synthesis of cDNA from template RNA (total RNA and mRNA), i.e., reverse transcription, is now possible. This is one of basic tools for genetic study and one of essential key techniques applied in various fields including molecular biology, biology, pharmaceuticals, forensic medicine, viral diagnosis, and the like.
In the conventional reverse transcription method, cDNA is synthesized at a relatively low temperature of 37° C. to 42° C. As a result, the template forms the secondary structure, which interrupts cDNA synthesis during reverse transcription reaction. To overcome this problem, a method using a thermophilic reverse transcriptase has been proposed. However, primers for reverse transcription have a short length, that is, random primers have the length of 5mer to 15mer and dT primers have the length of 14mer to 23mer. Therefore, at a high temperature, the primers could not be fully annealed with the template, resulting in decrease in the reaction efficiency.